Cationic polymer vehicles have emerged as promising platforms for nucleic acid delivery owing to their scalability, biocompatibility, and chemical versatility. Advancements in synthetic polymer chemistry allow us to precisely tune chemical functionality with various macromolecular architectures in order to increase the efficacy of nonviral based gene delivery. Herein, we demonstrate the first cationic bottlebrush polymer-mediated pDNA delivery by comparing unimolecular, synthetically-defined bottlebrush polymers to their linear building blocks. We successfully synthesized poly 2-(dimethylamino) ethyl methacrylate (pDMAEMA) bottlebrushes through ring opening metathesis polymerization to afford four bottlebrush polymers with systematic increases in backbone degree of polymerization (N_bb= 13, 20, 26, 37), while keeping the side-chain degree of polymerization constant (N_sc= 57). Physical and chemical properties were characterized, and subsequently, the toxicity and delivery efficiency of pDNA into HEK293 cells were evaluated. The pDNA complex (bottleplex) with the highest N_bb, BB_37, displayed up to a 60-fold increase in %EGFP+ cells in comparison to linear macromonomer. Additionally, we observed a trend of increasing EGFP expression with increasing polymer molecular weight. Bottleplexes and polyplexes both displayed high pDNA internalization as measured via payload enumeration per cell, however, quantitative confocal analysis revealed that bottlebrushes were able to shuttle pDNA into and around the nucleus more successfully than pDNA delivered via linear analogs. Overall, a canonical cationic monomer, such as DMAEMA, synthesized in the form of cationic bottlebrush polymers proved to be far more efficient in functional pDNA delivery and expression than linear pDMAEMA. This work underscores the importance of architectural modifications and the potential of bottlebrushes to serve as effective biomacromolecule delivery vehicles.

Rishad Dalal

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